Variant in situ Sequencing (VIS-seq) is a platform for optically profiling thousands of transgenically expressed protein-coding variants simultaneously. VIS-seq comprises a cassette with a promoter expressing a circular RNA containing one or more barcodes that are sequenced in situ to reveal the identity of the variant expressed in each cell and a second promoter expressing the protein variant. We used VIS-seq to create morphological profiles comprising a large set of measurements of the intensity, distribution and shape of different markers for >3,000 variants of lamin A and PTEN from ~11.4 million cell images. Lamin A variants were expressed in U2OS cells and PTEN variants in either iPS cells or derived neurons. VIS-seq variant profiles may be explored at visseq.gs.washington.edu.